Active peptide, active peptide composition and application of active peptide composition in preparation of product with skin photoaging resistance

Abstract

The invention discloses an active peptide, an active peptide composition and application of the active peptide composition in preparation of a product with a skin photoaging resisting effect. The squid active peptide is selected from a squid cyclic peptide-I, a squid cyclic peptide-II or a squid cyclic peptide-III; the squid cyclic peptide-I has a cyclic-(His-Val-Trp-Leu) amino acid sequence; the squid cyclic peptide-II has a cyclic-(His-Ala-Trp-Thr) amino acid sequence, and the squid cyclic peptide-II has an amino acid sequence; and the squid cyclic peptide-III has a cyclic-(Leu-Val-His-Phe) amino acid sequence. The squid active peptide composition comprises a squid cyclic peptide-I, a squid cyclic peptide-II and a squid cyclic peptide-III. The squid active peptide and the squid active peptide composition disclosed by the invention have the effects of resisting skin injury, skin photoaging, skin inflammation and skin oxidation; therefore, the compound has important application value when being used as an active ingredient for preparing cosmetics, food, health care products or medicines with corresponding effects.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technology, and in particular to an active peptide, an active peptide composition and application thereof in preparing a product with anti-skin photoaging effect. Background Art

[0002] Skin photoaging is the damage caused by long-term exposure to sunlight. It is the result of the combined effects of natural aging and ultraviolet radiation. The main manifestations of skin photoaging include roughness, thickening, dryness, loose skin, deeper and thicker wrinkles in exposed areas (such as the face, neck, chest, back, and upper forearms), and even various benign or malignant tumors, such as solar keratosis, squamous cell carcinoma, and malignant melanoma.

[0003] Oxidative damage is the damage to cells caused by the failure of free radicals such as reactive oxygen species (ROS) produced by cells during metabolism to be removed in time. Ultraviolet radiation is one of the important factors causing oxidative damage to the skin. Ultraviolet rays can stimulate the oxidative stress response of skin cells, leading to increased ROS production, which in turn attacks biological macromolecules such as cell membranes, DNA, and proteins, causing damage to cell structure and function. Inflammation is an automatic defense response of the skin to irritants or pathogens. During the process of skin photoaging, ultraviolet radiation can induce skin inflammatory reactions. Inflammatory cells (such as macrophages, neutrophils, etc.) will also infiltrate into damaged skin areas, releasing mediators such as inflammatory factors and proteases, and further aggravating skin cell damage and aging.

[0004] Ultraviolet radiation is a common cause of skin photoaging, oxidative damage and inflammation. Ultraviolet rays can penetrate the surface of the skin, directly damage skin cells, and induce oxidative stress and inflammatory reactions. Oxidative damage is an important intermediate link between ultraviolet radiation and skin aging. Ultraviolet radiation leads to increased ROS production, which in turn causes oxidative damage to cells, damaging cell structure and function. Inflammation is a response of the skin to ultraviolet radiation and oxidative damage. Infiltration of inflammatory cells and the release of inflammatory factors will further aggravate the damage and aging of skin cells.

[0005] As an important part of marine biological resources, squid contains abundant active substances such as proteins and peptides. These substances have various physiological functions, such as antibacterial, antioxidant, hypotensive, anti-tumor, etc., so they are widely used in food, medicine and health care products. For example, squid active peptides have inhibitory effects on a variety of bacteria such as Gram-positive bacteria and Gram-negative bacteria. Squid active peptides can remove free radicals in the body, reduce the damage of oxidative stress to cells, and thus play an antioxidant role. Some squid active peptides have the activity of inhibiting angiotensin converting enzyme (ACE) and play a role in lowering blood pressure. In recent years, studies have also found that some squid active peptides have an inhibitory effect on tumor cells. Squid active peptides can be used as functional food additives to improve the antioxidant, antibacterial and nutritional value of food. Due to its multiple biological activities, squid active peptides are of great value in the development of antibacterial drugs, antioxidants, antihypertensive drugs and anti-tumor drugs. Squid active peptides can also be used to develop health products with health functions such as antioxidant, anti-aging and immunity enhancement. However, there are few research reports and applications on the function and potential mechanism of squid active peptides in resisting skin damage and skin photoaging caused by ultraviolet light exposure. Summary of the invention

[0006] In order to overcome at least one of the technical problems existing in the prior art, the present invention provides an active peptide, an active peptide composition and the use thereof in preparing a product having an anti-skin photoaging effect.

[0007] The present invention provides a squid active peptide, wherein the squid active peptide is selected from squid cyclic peptide-I, squid cyclic peptide-II or squid cyclic peptide-III;

[0008] The squid cyclic peptide-I has a cyclic-(His-Val-Trp-Leu) amino acid sequence;

[0009] The squid cyclopeptide-II has a cyclo-(His-Ala-Trp-Thr) amino acid sequence;

[0010] The squid cyclic peptide-III has a cyclic-(Leu-Val-His-Phe) amino acid sequence.

[0011] The present invention provides a squid active peptide, wherein the squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III are prepared by the following method:

[0012] (1) mixing the squid with water, homogenizing and ultrasonically extracting the squid, centrifuging the extract and collecting the supernatant, and freeze-drying the supernatant to obtain freeze-dried powder;

[0013] (2) dialyzing the lyophilized powder through a dialysis membrane to obtain a dialyzed active fraction;

[0014] (3) separating the active dialyzed fraction by Sephadex G-15 column chromatography to obtain column chromatography active fractions;

[0015] (4) The column chromatography active fractions are separated by high performance liquid chromatography to obtain squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III, respectively.

[0016] The present invention also provides a method for preparing the above-mentioned squid active peptide, which comprises the following steps:

[0017] (1) mixing the squid with water, homogenizing and ultrasonically extracting the squid, centrifuging the extract and collecting the supernatant, and freeze-drying the supernatant to obtain freeze-dried powder;

[0018] (2) dialyzing the lyophilized powder through a dialysis membrane to obtain a dialyzed active fraction;

[0019] (3) separating the active dialyzed fraction by Sephadex G-15 column chromatography to obtain column chromatography active fractions;

[0020] (4) The column chromatography active fractions are separated by high performance liquid chromatography to obtain squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III, respectively.

[0021] The present invention also provides a squid active peptide composition, which comprises the squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III according to claim 1 or 2.

[0022] Preferably, the mass ratio of squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III is 1:(1-150):(1-150).

[0023] More preferably, the mass ratio of squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III is 1:(1-50):(1-50).

[0024] Most preferably, the optimal mass ratio of squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III is 1:5.8:8.1.

[0025] The present invention also provides an application of the squid active peptide or the squid active peptide composition in preparing a product with anti-skin damage effect.

[0026] Preferably, the anti-skin damage effect specifically refers to the effect of resisting skin damage caused by ultraviolet light exposure.

[0027] The present invention also provides an application of the squid active peptide or the squid active peptide composition in preparing a product with anti-skin aging effect.

[0028] Preferably, the anti-skin aging specifically refers to anti-skin photoaging caused by ultraviolet radiation.

[0029] The present invention also provides an application of the squid active peptide or the squid active peptide composition in preparing a product with anti-inflammatory effect.

[0030] Preferably, the anti-inflammatory effect specifically refers to anti-skin inflammation caused by ultraviolet light irradiation.

[0031] The present invention also provides an application of the squid active peptide or the squid active peptide composition in preparing a product with an antioxidant effect.

[0032] Preferably, the anti-oxidation specifically refers to resistance to skin oxidation caused by ultraviolet radiation.

[0033] Preferably, the product is a cosmetic, a food, a health product or a medicine.

[0034] Beneficial effects: The present invention provides a new squid active peptide and a squid active peptide composition; specific experiments show that the squid active peptide and the squid active peptide composition have the following effects:

[0035] The squid active peptide composition provided by the present invention can inhibit the increase of wrinkles on the skin of nude mice caused by ultraviolet light exposure; inhibit the peeling of the skin of nude mice caused by ultraviolet light exposure; inhibit the thickening of the stratum corneum of the skin of nude mice caused by ultraviolet light exposure, and exert anti-photoaging activity.

[0036] The squid active peptide composition provided by the present invention can inhibit the decrease in the activity of antioxidant enzymes (superoxide dismutase, SOD) in the skin tissue of nude mice caused by ultraviolet light exposure, inhibit the increase in the content of reactive oxygen species (ROS) in the skin tissue cells of nude mice caused by ultraviolet light exposure, and exert antioxidant activity.

[0037] The squid active peptide composition provided by the present invention can inhibit the increase in the content of inflammatory factors (TNF-α, IL-1β and IL-6) in skin tissue cells of nude mice caused by ultraviolet light exposure, and exert anti-inflammatory activity.

[0038] In summary, since the squid active peptide and the squid active peptide composition described in the present invention have anti-skin damage, anti-skin photoaging, anti-skin inflammation and anti-skin oxidation effects; therefore, using them as active ingredients for preparing cosmetics, foods, health products or medicines with corresponding effects has important application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 These are high-resolution mass spectra of squid cyclic peptide-I, squid cyclic peptide-II, and squid cyclic peptide-III.

[0040] Figure 2These are high-resolution mass spectra of squid cyclic peptide-I, squid cyclic peptide-II, and squid cyclic peptide-III.

[0041] Figure 3 These are high-resolution mass spectra of squid cyclic peptide-I, squid cyclic peptide-II, and squid cyclic peptide-III.

[0042] Figure 4 This is the experimental result of squid active peptide composition alleviating photodamage and photoaging of mouse skin.

[0043] Figure 5 This is the result of HE staining experiment on nude mouse skin sections using a squid active peptide composition to alleviate photodamage and photoaging of mouse skin.

[0044] Figure 6 This is the experimental result of squid active peptide composition alleviating photodamage and photoaging of mouse skin, oxidative and inflammatory damage of nude mouse skin. DETAILED DESCRIPTION

[0045] The present invention is further explained below in conjunction with specific examples, but the examples do not limit the present invention in any form.

[0046] Example 1 Preparation method of squid active peptide

[0047] (1) 2 kg of squid was mixed with 3 L of water, homogenized, and extracted by ultrasonic for 30 min. The extract was centrifuged for 10 min, and the supernatant was collected and freeze-dried to obtain 389.5 g of freeze-dried powder.

[0048] (2) 150 g of the lyophilized powder was dissolved in 1 L of water, and then dialyzed using a 1 kDa dialysis membrane. The dialysate was lyophilized to obtain 18.2 g of the dialyzed active fraction;

[0049] (3) 18.2 g of the dialyzed active fraction was dissolved in 50 mL of water, and then chromatographed on a Sephadex G-15 column (specification: 112*3.5 cm), followed by elution with a 15% ethanol aqueous solution; from the start of elution, each 1 / 2 column volume of the eluate was collected as a fraction; 5 fractions were collected continuously; the second fraction was concentrated to remove ethanol and freeze-dried to obtain 2.1 g of the column chromatography active fraction;

[0050] (4) taking the active fraction from the column chromatography and separating it by high performance liquid chromatography to obtain squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III respectively;

[0051] The specific conditions of the high performance liquid chromatography are as follows: the chromatographic column is a C18 column (4.6×250 mm, 5 μm, Waters, USA); the mobile phase A is water, and the mobile phase B is methanol; the elution method is gradient elution, wherein the gradient elution program is: 0-10 min 25% B, 11-22 min 55% B; the injection volume is 100 μL; the detection wavelength is 230 nm; the flow rate is 2.0 mL / min; and the column temperature is 18°C;

[0052] The eluate corresponding to the chromatographic peak with a retention time of 8.24 min was collected, concentrated and freeze-dried to obtain squid cyclic peptide-I;

[0053] The eluate corresponding to the chromatographic peak with a retention time of 9.14 min was collected, concentrated and freeze-dried to obtain squid cyclic peptide-II;

[0054] The eluate corresponding to the chromatographic peak with a retention time of 7.44 min was collected, concentrated and freeze-dried to obtain squid cyclic peptide-III.

[0055] The structural analysis of squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III is as follows:

[0056] Squid cyclopeptide-I: Analytical ESI-MS data, m / z 536.2996 is [M+H] + Ion; m / z 437.2300 is [M-Val+H2O+H] + Ion; m / z 423.2135 is [M-Leu+H2O+H] + ion; m / z 300.1709 is [Trp-Leu-2H2O+H] + ion; m / z 251.1502 is [Leu-His-2H2O+H] + ion; m / z 159.0913 is [Trp-COOH+H] + ion; m / z 110.0717 is [His-COOH+H] + ion; m / z 86.0970 is [Leu-COOH+H] + Based on the mass spectrometry data and the Marfey method analysis experimental results, the compound was finally identified as a cyclic tetrapeptide with a cyclo-(His-Val-Trp-Leu) amino acid sequence, which was named squid cyclic peptide-I in the present invention.

[0057] Squid cyclopeptide-II: Analyzing ESI-MS data, m / z 496.7325 is [M+H] + ion; m / z 478.2213 is [M-H2O+H] + Ion; m / z 460.7127 is [M-2H2O+H]+ Ion; m / z 425.1942 is [M-Ala+H2O+H] + ion; m / z 258.1244 is [Trp-Ala-2H2O+H] + ion; m / z 239.1140 is [His-Thr-2H2O+H] + ion; m / z 209.1035 is [Ala-His-2H2O+H] + ion; m / z 159.0920 is [Trp-COOH+H] + ion; m / z 110.0721 is [His-COOH+H] + Based on the mass spectrometry data and Marfey method analysis experimental results, the compound was finally identified as a cyclic tetrapeptide with a cyclo-(His-Ala-Trp-Thr) amino acid sequence, which was named squid cyclic peptide-II in the present invention.

[0058] Squid cyclopeptide-III: Analytical ESI-MS data, m / z 497.2843 is [M+H] + ion; m / z 469.2897 is [M-CO+H] + Ion; m / z 398.2164 is [M-Val+H2O+H] + ion; m / z 384.2012 is [M-Leu+H2O+H] + Ion; m / z 380.2053 is [M-Val+H] + ion; m / z 350.2168 is [M-Phe+H2O+H] + ion; m / z 285.1325 is [His-Phe-2H2O+H] + ion; m / z 261.1582 is [Leu-Phe-2H2O+H] + ion; m / z 237.1332 is [His-Val-2H2O+H] + Ion; m / z 212.1169 is [Val-Leu-2H2O+H] + ion; m / z 138.0656 is [His-H2O+H] + ion; m / z 110.0708 is [His-COOH+H] + ion; m / z 86.0975 is [Leu-COOH+H] +Based on the mass spectrometry data and the Marfey method analysis experimental results, the compound was finally identified as a cyclic tetrapeptide with a cyclic-(Leu-Val-His-Phe) amino acid sequence, which was named squid cyclic peptide-III in the present invention.

[0059] Example 2 Preparation of squid active peptide composition

[0060] The squid cyclic peptide-I prepared according to the method of Example 1 is evenly mixed with the squid cyclic peptide-II and the squid cyclic peptide-III at a mass ratio of 1:5.8:8.1 to obtain the squid active peptide composition with the function of resisting skin photodamage and photoaging according to the present invention.

[0061] Example 3 Experimental evaluation of squid active peptide composition against photodamage and photoaging of nude mice skin

[0062] The squid active peptide composition in this experimental example refers to the squid active peptide composition prepared according to the method described in Example 2; the commercially available squid peptide used is a water-soluble small molecule squid peptide of ≤500da produced by Blue Power Biotechnology (Xi'an) Co., Ltd.

[0063] Twenty adult male nude mice (23±3g) were raised for one week and randomly divided into four groups, each with five mice, namely, a blank group, a light-damaged group, a commercially available squid peptide group, and a squid active peptide composition group of the present invention. The blank group was not treated and the back skin was kept exposed; the skin light-damaged group was treated with an ultraviolet lamp [ultraviolet intensity: UVA (365nm) = 300uW·cm -2 and UVB (310nm) = 30uw·cm -2 ] Irradiate the back skin of mice daily. Irradiate for 3 min / d in the first 1-2 weeks, and then increase by 20 s per week for a total of 5 weeks; the commercially available squid peptide group and the squid active peptide composition group of the present invention: apply the skin of mice after irradiating the skin with ultraviolet rays for 30 min (the final concentration of the commercially available squid peptide and the squid active peptide composition of the present invention is 1 mg / mL). Observe the behavioral characteristics of the animals every day, anesthetize and kill the mice 12 hours after the last application of the active substance, and collect the skin tissue for morphological observation and biochemical determination.

[0064] The experimental results show that ( Figure 4 ), the skin structure of the nude mice in the blank group was intact and undamaged; the skin of the nude mice in the photodamage group showed obvious damage, with obvious skin peeling and wrinkling, indicating that ultraviolet irradiation significantly damaged the skin of the nude mice and caused skin aging. Compared with the photodamage group, the squid active peptide composition of the present invention alleviated the skin damage and photoaging of nude mice caused by ultraviolet irradiation; compared with the commercially available squid peptide group of the same concentration, the squid active peptide composition of the present invention alleviated the skin damage of nude mice caused by ultraviolet irradiation, and alleviated the photoaging of nude mice caused by ultraviolet irradiation.

[0065] Analysis of the results of HE staining of nude mouse skin tissue ( Figure 5 ) showed that ultraviolet irradiation caused thickening of the stratum corneum and damage to skin cells of nude mice; the squid active peptide combination of the present invention can effectively alleviate the thickening of the stratum corneum of nude mice caused by ultraviolet irradiation, and reduce the damage to dermal cells of nude mice skin caused by ultraviolet irradiation; and compared with the same concentration of commercially available squid peptides, the squid active peptide combination of the present invention can more effectively alleviate the thickening of the stratum corneum of nude mice skin caused by ultraviolet irradiation, and can more effectively reduce the damage to dermal cells of nude mice skin caused by ultraviolet irradiation, and exert protective activity.

[0066] Skin photodamage refers to damage caused by exposure to natural light sources (such as sunlight) or artificial light sources (such as ultraviolet lamps, lasers, etc.) to the skin. This damage mainly comes from ultraviolet radiation, especially the two ultraviolet bands of UVA and UVB. Ultraviolet radiation can directly or indirectly damage DNA, affect the normal function of cells, and even cause the occurrence of skin tumors. Ultraviolet radiation can also cause inflammatory reactions in the skin, causing the skin to become thinner and brittle, and accelerate skin aging. Ultraviolet radiation and the oxidative damage and DNA damage it causes can lead to cell apoptosis, further aggravating skin damage. Among them, oxidative damage refers to the imbalance in the removal of oxygen free radicals in the body, producing a large number of highly active reactive oxygen species (ROS), which will attack biological macromolecules such as cell membranes, proteins, and DNA, leading to damage to cell structure and function.

[0067] Analyze the experimental results ( Figure 6A) It was found that compared with the blank group (the ROS content in the skin tissue cells of the blank group was set to 100%, and the antioxidant enzyme SOD activity in the skin tissue cells was set to 100%), the ROS content in the skin tissue cells of the nude mice in the light damage group increased by 226.3% (p<0.01), and the antioxidant enzyme SOD activity decreased by 35.9% (p<0.01). We speculate that ultraviolet irradiation induces oxidative stress in the skin of nude mice and causes severe oxidative damage to the skin cells of nude mice. Compared with the skin photodamage group, after treatment with the squid active peptide composition of the present invention, the increase in ROS content in the skin tissue cells of nude mice induced by ultraviolet irradiation and the decrease in the activity of the antioxidant enzyme SOD were significantly reversed. Compared with the light damage group, the ROS content in the skin tissue cells of the commercially available squid peptide and the squid active peptide composition group of the present invention decreased by 9.20% and 42.0% respectively (p<0.01); and compared with the light damage group, the activity of the antioxidant enzyme SOD in the skin tissue cells of nude mice in the commercially available squid peptide and the squid active peptide composition group of the present invention increased by 9.59% and 30.9% respectively (p<0.01). Further analysis of the experimental results showed that compared with the same concentration of commercially available squid peptides, the squid active peptide composition of the present invention has a stronger ability to inhibit the increase of ROS content in the skin tissue cells of nude mice caused by ultraviolet irradiation, and improve the activity of the antioxidant enzyme SOD in the skin tissue cells of nude mice caused by ultraviolet irradiation, and has better antioxidant activity.

[0068] Inflammation is a defensive response of the body to stimulation, manifested as redness, swelling, heat, pain and dysfunction, etc. In the process of skin photoaging, inflammation is an important pathophysiological process, and inflammatory factors are a type of cytokines that participate in and mediate inflammatory responses. They are produced by immune cells (such as macrophages, T cells, etc.) or other types of cells (such as endothelial cells, epidermal cells, etc.) under stress conditions.

[0069] In skin photoaging, ultraviolet radiation activates immune cells in the skin, leading to an inflammatory response. These immune cells release inflammatory factors, such as IL-1 and TNFα, which further aggravate the inflammatory response of the skin. In addition, inflammatory factors also promote the secretion of enzymes such as matrix metalloproteinases (MMPs), which degrade extracellular matrix components in the skin, such as collagen and elastic fibers, leading to skin structural damage and aging. At the same time, the inflammatory response caused by ultraviolet radiation accelerates the apoptosis of skin cells and the degradation of the extracellular matrix, leading to accelerated skin aging.

[0070] The results of the experiment on the effect of the squid active peptide composition of the present invention on the excessive secretion of inflammatory cytokines in the skin tissue of nude mice induced by ultraviolet irradiation ( Figure 6C, D, E) show. Compared with the blank group (the contents of IL-6, IL-1β and TNF-α in the skin tissue of the blank group were set as 100%), the contents of inflammatory cytokines IL-6, IL-1β and TNF-α in the skin tissue of nude mice in the photodamage group increased by 258.6% (p<0.01), 172.8% (p<0.01) and 155.3% (p<0.01), respectively. This shows that ultraviolet irradiation induces increased secretion of inflammatory cytokines in the skin tissue of nude mice, aggravates the inflammatory response and further damages the skin tissue, making the photoaging of the skin of nude mice more serious. After treatment with the squid active peptide composition of the present invention, the excessive inflammatory cytokines in the skin tissue of nude mice caused by ultraviolet irradiation were effectively inhibited. Compared with the light damage group, the contents of inflammatory cytokines IL-6, IL-1β and TNF-α in the skin tissue of nude mice in the squid active peptide composition group of the present invention were reduced by 28.8% (p<0.01), 30.9% (p<0.01) and 33.7% (p<0.01), respectively. And compared with the commercially available squid peptides, the squid active peptide composition of the present invention at the same concentration can better inhibit the excessive secretion of inflammatory cytokines in the skin tissue of nude mice caused by ultraviolet irradiation, exert a better anti-inflammatory activity, and then exert a better protective activity, reducing the light damage and photoaging of the skin of nude mice caused by ultraviolet irradiation.

[0071] Photoaging refers to the process by which long-term exposure to ultraviolet radiation causes skin damage and aging. Ultraviolet radiation activates immune cells in the skin, such as macrophages and T cells, which in turn triggers an inflammatory response. This inflammatory response releases inflammatory factors, such as cytokines and chemokines, which further damage skin cells and accelerate the skin aging process. Specifically, UVB radiation in ultraviolet rays can directly damage the DNA of skin cells, while UVA radiation can activate metalloenzymes to degrade elastic fibers and collagen, while producing a large number of free radicals, leading to oxidative stress and cell damage. These processes will aggravate the inflammatory response and accelerate skin aging.

[0072] The NLRP3 inflammasome is a complex composed of multiple proteins, including NLRP3 (NOD-like receptor protein 3), ASC (apoptosis-associated microparticle protein, containing CARD and PYD domains) and Caspase-1 (caspase-1). When NLRP3 is activated, it binds to ASC and Caspase-1 to form an oligomeric complex, which is called the NLRP3 inflammasome. As an important component of innate immunity, the NLRP3 inflammasome plays an important role in the body's immune response and the occurrence of diseases. It can aggregate in response to cell perturbations, leading to the activation of caspase-1, which in turn promotes the maturation and release of inflammatory cytokines interleukin 1β (IL-1β) and IL-18, as well as inflammatory cell death (pyroptosis). These inflammatory cytokines contribute to the development of systemic low-grade inflammation, and abnormal NLRP3 activation can drive the chronic inflammatory state in the body, thereby regulating the pathogenesis of a variety of inflammatory-related diseases. At present, there are few studies on the relationship between photoaging and the NLRP3 inflammasome, but the analysis of experimental results ( Figure 6 F) found that compared with the blank group (the NLRP3, ASC and Caspase-1 protein contents in the skin tissue of the blank group were all set to 100%), the NLRP3, ASC and Caspase-1 protein contents in the skin tissue of the nude mice in the photodamage group increased by 199.6% (p<0.01), 201.5% (p<0.01) and 89.3% (p<0.01), respectively. It shows that ultraviolet irradiation induces increased secretion of inflammatory cytokines in the skin tissue of nude mice and also activates the NLRP3 inflammasome, aggravating the inflammatory response. After treatment with the squid active peptide composition of the present invention, the activation of the NLRP3 inflammasome in the skin tissue of nude mice caused by ultraviolet irradiation was effectively inhibited. Compared with the photodamage group, the NLRP3, ASC and Caspase-1 protein contents in the skin tissue of nude mice in the squid active peptide combination group of the present invention decreased by 48.1% (p<0.01), 44.9% (p<0.01) and 32.8% (p<0.01), respectively. Compared with commercially available squid peptides, the squid active peptide composition of the present invention at the same concentration can better inhibit the activation of NLRP3 inflammasomes in skin tissue cells of nude mice caused by ultraviolet light irradiation and exert better anti-inflammatory activity, thereby exerting better anti-photodamage and anti-photoaging activity of nude mice skin.

[0073] Studies have found that the inflammatory response caused by ultraviolet irradiation can further aggravate the damage and aging of skin cells by activating the NLRP3 inflammasome. For example, ultraviolet irradiation may trigger the activation of the NLRP3 inflammasome by producing ROS (reactive oxygen species) or other intracellular signals, thereby promoting the release of inflammatory cytokines such as IL-1β and IL-18, which will further aggravate the inflammatory response and damage of skin cells. Compared with the commercially available squid peptides, the squid active peptide composition of the present invention can better inhibit the excessive accumulation of ROS and the reduction of SOD enzyme activity in the skin tissue cells of nude mice caused by ultraviolet irradiation, and can better inhibit the excessive secretion of inflammatory cytokines in the skin tissue cells of nude mice caused by ultraviolet irradiation. At the same time, it can also better inhibit the activation of the NLRP3 inflammasome of the skin tissue cells of nude mice caused by ultraviolet irradiation, and exert better anti-inflammatory activity and antioxidant activity, and exert better anti-nude mouse skin light damage and anti-photoaging activity.

Claims

1. A squid active peptide, characterized in that: The squid active peptide is selected from squid cyclic peptide-I, squid cyclic peptide-II or squid cyclic peptide-III; The squid cyclic peptide-I has a cyclic-(His-Val-Trp-Leu) amino acid sequence; The squid cyclopeptide-II has a cyclo-(His-Ala-Trp-Thr) amino acid sequence; The squid cyclic peptide-III has a cyclic-(Leu-Val-His-Phe) amino acid sequence.

2. The squid active peptide according to claim 1, characterized in that: The squid cyclic peptide-I and squid cyclic peptide -II and squid cyclic peptide-III were prepared by the following method: (1) mixing the squid with water, homogenizing and ultrasonically extracting the squid, centrifuging the extract and collecting the supernatant, and freeze-drying the supernatant to obtain freeze-dried powder; (2) dialyzing the lyophilized powder through a dialysis membrane to obtain a dialyzed active fraction; (3) Separating the active dialysis fraction by Sephadex G-15 column chromatography, Column layer Analysis of active sites; (4) Separating the column chromatography active fractions by high performance liquid chromatography to obtain squid cyclic peptide-I and squid cyclic peptide, respectively -II and squid cyclopeptide-III.

3. A method for preparing squid active peptides, characterized in that: The following steps are included: (1) mixing the squid with water, homogenizing and ultrasonically extracting the squid, centrifuging the extract and collecting the supernatant, and freeze-drying the supernatant to obtain freeze-dried powder; (2) dialyzing the lyophilized powder through a dialysis membrane to obtain a dialyzed active fraction; (3) Separating the active dialysis fraction by Sephadex G-15 column chromatography, Column layer Analysis of active sites; (4) Separating the column chromatography active fractions by high performance liquid chromatography to obtain squid cyclic peptide-I and squid cyclic peptide, respectively -II and squid cyclopeptide-III.

4. A squid active peptide composition, characterized in that: It comprises the squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III as described in claim 1 or 2.

5. The squid active peptide composition according to claim 3, characterized in that: The mass ratio of squid cyclic peptide-I, squid cyclic peptide-II and squid cyclic peptide-III is 1:(1-150):(1-150).

6. Use of the squid active peptide or squid active peptide composition according to any one of claims 1 to 5 in the preparation of products having anti-skin damage effects.

7. Use of the squid active peptide or squid active peptide composition according to any one of claims 1 to 5 in the preparation of products having anti-skin aging effects.

8. Use of the squid active peptide or squid active peptide composition according to any one of claims 1 to 5 in the preparation of products with anti-inflammatory effects.

9. Use of the squid active peptide or squid active peptide composition according to any one of claims 1 to 5 in the preparation of products with antioxidant effects.

10. The use according to any one of claims 6 to 9, characterized in that: The product is a cosmetic, food, health product or medicine.

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